Abstract Modeling sex differences in Alzheimer?s Disease cognition and pathology There are significant differences between men and women in the incidence and prevalence of Alzheimer?s Disease (AD). After menopause, women are more likely to develop AD, and symptoms of the disease including cognitive impairment are more severe. These sex differences are further complicated by high cholesterol which, at midlife, is a major risk factor for AD, and there is substantial interaction between estrogen and cholesterol. There is a significant gap in our knowledge of how estrogen neuroprotection and cholesterol diet-associated vulnerability converge because replacing estrogen or treating with cholesterol-lowering statins may not reverse cognitive impairment and can even make it worse. We propose to model sex differences in AD and the complicating effects of high cholesterol by studying cholesterol-fed male and female rabbits because they show significant sex differences in AD-like pathology and cognition. Cholesterol-fed females develop beta amyloid (A?) deposits more slowly than cholesterol-fed males and eliminating peripheral estrogen by ovariectomy more than doubles A? levels, suggesting a protective role for estrogen against amyloid pathology. We have preliminary data suggesting female cholesterol-fed rabbits remember trace eyeblink conditioning better than cholesterol-fed males and that a cholesterol diet alters estrogen receptors alpha and beta which correlates with a significant increase in serum and hippocampal levels of the cholesterol metabolite, 27-hydroxycholesterol (27- OHC). 27-OHC is an endogenous estrogen receptor modulator that may play a role in learning and memory because patients with mild cognitive impairment (MCI) and AD exhibit elevated 27-OHC levels and we have preliminary data suggesting cholesterol-fed rabbits with elevated 27-OHC have memory deficits. We also have new data showing sex differences in the transcriptional activity of estrogen receptors and expression of proteins in the presynaptic active zone and postsynaptic density that are higher in female cholesterol-fed rabbits than males. The present research focus on cholesterol-induced increases in 27-OHC has direct clinical relevance because midlife hypercholesterolemia is a risk factor for AD and, importantly, 27-OHC is significantly elevated in mild cognitive impairment and AD. In two specific aims, we will manipulate estrogen (Aim 1) and estrogen receptors (Aim 2) in cholesterol-fed rabbits to test the hypothesis that sex differences in AD-like cognitive impairment and pathology are a function of endogenous estrogen receptor modulation of downstream targets. Using behavioral, electrophysiological, histochemical and molecular biological techniques, we will determine the mechanisms by which endogenous estrogen receptor modulation by cholesterol diet-induced 27-OHC affects memory, neural function, markers of cholesterol and A? processing, and A? and tau levels in male and female rabbits. Our combined expertise in and track record of behavioral, histochemical, electrophysiological and molecular biological research in cholesterol-fed rabbits makes us a particularly well-suited collaborative team to conduct these experiments and places us in a strong position to have a significant impact on the field.